Schizophrenia is a mental disorder that affects approximately 1% of the general population. The disease is clinically complex and heterogeneous with respect to symptomatology, disease course and responses to treatment. It is a very disabling condition, often striking in early adult years and resulting in long-term drug therapy, hospitalisation and in many cases an inability to pursue a normal lifestyle and gainful employment. Characteristic symptoms include psychotic episodes, such as hallucinations and delusions; thought disorder; disorganised behaviour and negative symptoms such as social withdrawal; diminished emotional range and poverty of speech. Treatment strategies are usually a combination of neuroleptic medication and psychosocial therapy. First generation neuroleptics were developed serendipitously and have been associated with significant side effects and low efficacy. Although new generation neuroleptics are providing some improved disease management options for a larger proportion of individuals with schizophrenia, to date there is no cure for schizophrenia and the risk factors and pathology of the disease are unknown.

An extensive body of epidemiological research has demonstrated that susceptibility to schizophrenia can be attributed largely to heritable factors. The favoured model proposes the interplay of genetic and environmental factors to create a state of increased risk for schizophrenia. Family-based studies have consistently suggested a significant heritability component of schizophrenia that does not follow known Mendelian inheritance patterns. It is now widely accepted that schizophrenia susceptibility is genetically complex, involving multiple genes of small to moderate effect, each with varying degrees of penetrance amongst the affected populations. This complexity has hampered research efforts to identify the genetic elements involved. With respect to environmental factors, some evidence suggests that factors such as season of birth (winter/spring births), obstetric complications, urban versus rural childhood and viral exposure act to increase the risk to develop schizophrenia later in life. A clear understanding of the environmental factors involved in schizophrenia and their relationship to genetic elements is yet to be determined.

To date, research efforts towards understanding the genetic mechanisms involved in schizophrenia have relied heavily upon two approaches, 1) genome-wide screening using genetic linkage analysis and 2) candidate gene analysis using mutation screening techniques. Genome scan methodologies and experimental designs have undergone several developments during the course of their application to complex disease genetics. As a result, recent years have seen several large collaborative efforts replicate previous findings of chromosomal regions providing suggestive evidence of linkage to schizophrenia linkage. The regions implicated include 6p22- 24, 10p11-p15, 22q11-q12, 6q13-q26, Iq32-q42 and 13q14.1-q32. To date, there are no replicated findings of linkage that satisfy statistical criteria for confirmed linkage as proposed by Lander and Kruglyak (1995).

Candidate gene studies have focussed upon genes involved in the neurotransmitter pathways implicated by neuroleptic pharmacology and psychotic mimicry by recreational drugs. Several associations have been reported, however, to date no findings have proven to be robust across different clinical samples. This thesis describes the application of three independent approaches using both new and established experimental methods to the identification of genes and gene pathways involved in schizophrenia susceptibility.

The first approach characterised the contribution of the 22q11.2 microdeletion region to schizophrenia susceptibility in a collection of Australian and North American multiplex schizophrenia pedigrees. The 22q11.2 microdeletion has been implicated in 22q11 Deletion Syndromes (22q11DS). Facial dysmorphologies typical of 22q11DS have been observed in a subset of individuals with schizophrenia, and the 22q11DS have been shown to incorporate an increased risk of psychiatric symptomatology, in particular paranoid schizophrenia. The connection between the 22q11.2 microdeletion and schizophrenia has been further supported by findings of a region of suggestive linkage to 22q11-q13.

The work presented in this thesis represents the first investigation of the incidence of the 22q11.2 deletion in multiplex schizophrenia families. At the time of completion, this work was also the first investigation of the incidence of the 22q11.2 deletion in healthy population controls rather than unaffected relatives of 22q11DS patients. No evidence of an increased abundance of 22q11.2 deletions was detected in affected individuals. Similarly, no evidence was detected in multiplex schizophrenia pedigrees of segregation of the 22q11.2 deletion with schizophrenia susceptibility. These findings indicate deletion of 22q11.2 does not contribute significantly to schizophrenia linkage in the 22q11-13 region and suggests that further investigation of linkage finding in this region is required.

The second approach used a mutation screening methodology, (denaturing gel electrophoresis) in a high-throughput experimental design to investigate gene candidates within the region of the suggested linkage on 6q13-26. Two serotonin receptor genes, HTR1B and HTR1E map to this region. Serotonin neurotransmission is a strong component of several models for the pathology of schizophrenia; hence, within this region of suggested linkage the serotonin receptor genes are the obvious candidates. The work presented in this thesis represents the most comprehensive mutation screen of the coding regions of these genes conducted to date. Within the HTR1E and HTR1B genes respectively, five and four novel mutations were described. A large sample of 183 Australian and North American schizophrenia pedigrees was used to assess the contribution of these variants to schizophrenia susceptibility. While the analysis did not support the involvement of genetic variation of these genes in schizophrenia, it does demonstrate the screening procedure to be a fast, efficient method for screening candidate genes. The presented findings have substantial implications for further investigations in schizophrenia, particularly given the growing support for linkage to this 6q region.

The third approach was a preliminary investigation into the use of expression profiling to identify genes involved in schizophrenia. Expression profiling has gained immense popularity as a high-throughput screening method to identify genetic pathways associated with disease. Previous investigations have been able to detect growth and adhesion differences in fibroblast cultures from individuals with schizophrenia. An experimental design was developed to investigate the potential for fibroblast expression profiles to reflect these differences and discriminate between cultures from schizophrenic and control individuals. Since the commencement of this component of the thesis, three preliminary investigations of post-mortem brain samples have been published. Interestingly, some overlap exists between the findings from brain expression profiling studies and preliminary results from the fibroblast study presented here. Specifically, genetic dysregulation of membrane associated secondary messenger systems and GABAergic pathways is implicated. Further, the potential of fibroblast expression profiles to be able to discriminate between subgroups of schizophrenia and control samples is promising.

When used as tools, each experimental approach used in this thesis has specific advantages and limitations in the identification of genes for complex diseases. These methodologies represent a progression within the field of complex disease genetics from genome screen and linkage studies to high-throughput mutation screening and expression profiling approaches. These methodologies have begun to exploit the wealth of information being generated as a result of human genome sequencing projects and other large-scale genomic mapping projects. It is unlikely that a single methodological approach will be able to unravel the genetic complexity of schizophrenia susceptibility. Within this thesis, and in the wider schizophrenia genetic research field, it is acknowledged that the integration of information from a variety of research tools and approaches will be essential to achieve effective model generation, elucidate genetic and environmental risk factors, and ultimately lead to improved health outcomes for schizophrenia.